Ovarian Contractions Research Articles

Granulosa cell endothelin-2 expression is fundamental for ovulatory follicle rupture

Ovulation is dependent upon numerous factors mediating follicular growth, vascularization, and ultimately oocyte release via follicle rupture. Endothelin-2 (EDN2) is a potent vasoconstrictor that is transiently produced prior to follicle rupture by granulosa cells of periovulatory follicles and induces ovarian contraction. To determine the role of Edn2 expression, surgical transplant and novel conditional knockout mice were super-ovulated and analyzed. Conditional knockout mice utilized a new iCre driven by the Esr2 promoter to selectively remove Edn2. Follicle rupture and fertility were significantly impaired in the absence of ovarian Edn2 expression. When ovaries of Edn2KO mice were transplanted in wild type recipients, significantly more corpora lutea containing un-ovulated oocytes were present after hormonal stimulation (1.0 vs. 5.4, p = 0.010). Following selective ablation of Edn2 in granulosa cells, Esr2-Edn2KO dams had reduced oocytes ovulated (3.8 vs. 16.4 oocytes/ovary) and smaller litters (4.29 ± l.02 vs. 8.50 pups/dam). However, the number of pregnancies per pairing was not different and the reproductive axis remained intact. Esr2-Edn2KO ovaries had a higher percentage of antral follicles and fewer corpora lutea; follicles progressed to the antral stage but many were unable to rupture. Conditional loss of endothelin receptor A in granulosa cells also decreased ovulation but did not affect fecundity. These data demonstrate that EDN2-induced intraovarian contraction is a critical trigger of normal ovulation and subsequent fecundity.

Production and binding of endothelin-2 (EDN2) in the rat ovary: endothelin receptor subtype A (EDNRA)-mediated contraction

Endothelin-2 (EDN2)-mediated contraction has been proposed as a final mechanical signal facilitating ovulation. The objectives herein were to determine (1) whether ovarian endothelins were increased before ovulation; (2) whether a specific endothelin-converting enzyme (ECE) was mediating their production; (3) which receptor was facilitating ovarian contraction; and (4) whether receptor-specific antagonism affected ovulation. Follicular development was induced in immature rats with 10 IU pregnant mare serum gonadotrophin (PMSG) and the ovulatory cascade was initiated 48 h later with 10 IU human chorionic gonadotrophin (hCG). In Experiment 1, an immunoassay revealed that the ovarian concentration of endothelin peptide was increased 7-fold 12 h after hCG when compared with 48 h after PMSG (P < 0.05). In Experiment 2, real-time PCR indicated that mRNA for Ece1, but not Ece2, was increased in granulosa cells collected 12 h after hCG when compared with those collected before the ovulatory stimulus (P < 0.05). In Experiment 3, isometric tension analysis revealed that the contractile effect of EDN2 was mediated by endothelin receptor A (EDNRA), not B (EDNRB). In Experiment 4, no effect was observed on the rate of ovulation when rats were treated with an antagonist specific to EDNRA (BQ123) or EDNRB (BQ788), or when mice were treated with BQ123, BQ788 or BQ123 + BQ788. In conclusion, endothelin peptide is produced before ovulation and the contractile action of EDN2 within the ovary is facilitated via EDNRA. In addition, findings of this study indicate synergistic interactions among contractile factors affect ovulatory outcome, while the role of EDNRB alone in the process of ovulation requires further investigation.

Ovarian contraction in various teleost families

Fishes from 12 species (a minimum of three fish of each species), representing nine families were examined for ovarian contraction. An ovary from each fish was removed, placed in a muscle bath and assessed for contractions using a force‐displacement transducer. After at least 30 min of observation for spontaneous contractions, acetylcholine (ACh) was added to the muscle bath to assess the ovary’s capacity for contraction. At least one third of the ovaries from each species contracted spontaneously or in response to 10−5 M ACh. Mean ±s.e. frequencies of spontaneous contractions ranged from 7·5 ± 2·0 h−1 in the Atlantic silverside Menidia menidia to 380·3 ± 101·5 h−1 in the sheepshead minnow Cyprinodon variegatus. Simultaneously tested ovaries from a single white perch Morone americana contracted at rates of 31·4 and 81·9 h−1. This is consistent with independent generation of the contractile rhythm in the two ovaries. In three species (goldfish Carassius auratus, mummichog Fundulus heteroclitus and white perch), the tunica albuginea was removed from the ovary and found to be capable of independent contraction. This suggests that this membrane contributes to the observed contractions. The presence of ovarian contractions in this varied group of fish species indicates that the contractions play an important role in reproduction.

Vesicular Communication Between Sperm and Egg

Sperm and oocytes communicate and, in some organisms, such as Caenorhabditis elegans , sperm promote oocyte maturation through resumption of meiosis. Major sperm protein (MSP) serves a cytoskeletal role within spermatozoa and also serves as a hormone signaling oocyte meiotic maturation and ovarian muscle contraction in C. elegans . Kosinski et al. used immunofluorescence microscopy to show that MSP appears inside the spermatozoa and extracellularly in the proximal gonad arm (the site of the unfertilized oocytes) and in the uterus. Transmission electron microscopy (TEM), combined with immunohistochemistry, of samples prepared by high-pressure freezing and freeze substitution revealed the presence of a novel class of MSP-positive extracellular double-membrane vesicles in the spermatheca, in the uterine lumen, and between embryos. In wild-type hermaphrodites, oocyte maturation ceases and activation of mitogen-activated protein kinase (MAPK) decreases as sperm are depleted. MSP also becomes undetectable following sperm depletion. In mutant worms defective in spermiogenesis (they make spermatids, but these do not mature), oocyte maturation and MAPK activation persisted after spermatids were no longer present. MSP was also faintly detectable, suggesting that spermatids and spermatozoa release distinct MSP signals. Spermatid-released MSP is more stable than spermatozoa-released MSP. Thus, vesicle budding through a nontraditional secretory pathway (sperm lack endoplasmic reticulum and Golgi) appears to provide a mechanism of sperm-egg communication. M. Kosinski, K. McDonald, J. Schwartz, I. Yamamoto, D. Greenstein, C. elegans sperm bud vesicles to deliver a meiotic maturation signal to distant oocytes. Development 132 , 3357-3369 (2005). [Abstract] [Full Text]

Tropomyosin and troponin are required for ovarian contraction in the Caenorhabditis elegans reproductive system.

Ovulation in the nematode Caenorhabditis elegans is coordinated by interactions between the somatic gonad and germ cells. Myoepithelial sheath cells of the proximal ovary are smooth muscle-like cells, but the regulatory mechanism of their contraction is unknown. We show that contraction of the ovarian muscle requires tropomyosin and troponin, which are generally major actin-linked regulators of contraction of striated muscle. RNA interference of tropomyosin or troponin C caused sterility by inhibiting ovarian contraction that is required for expelling mature oocytes into the spermatheca where fertilization takes place, thus causing accumulation of endomitotic oocytes in the ovary. Tropomyosin and troponin C were associated with actin filaments in the myoepithelial sheath, and the association of troponin C with actin was dependent on tropomyosin. A mutation in the actin depolymerizing factor/cofilin gene suppressed the ovulation defects by RNA interference of tropomyosin or troponin C. These results strongly suggest that tropomyosin and troponin are the actin-linked regulators for contraction of ovarian muscle in the C. elegans reproductive system.

Effects of ions and bioactive substances on ovarian contraction in Fundulus heteroclitus

AbstractThe effects of potassium, calcium, angiotensin II, urotensin II, melatonin, gonadotropin releasing hormone analogue (GnRH‐a), and human chorionic gonadotropin (hCG) on in vitro ovarian smooth muscle contraction were observed in Fundulus heteroclitus. Calcium induced contractions and modified spontaneous contractions in a dose‐dependent manner. Potassium, angiotensin II and urotension II induced dose‐dependent contractions while melatonin, GnRH‐a and HCG had no observable effect on the system. These data highlight the integral role of calicum in ovarian contraction and suggest possible roles in reproduction for angiotensin II and urotensin II. © 1993 wiley‐Liss, Inc.

Spontaneous ovarian contractions in Fundulus heteroclitus

AbstractSpontaneous contractions of Fundulus heteroclitus ovaries were observed and quantified using 2 methods. Contractions were measured as increases in intraovarian pressure in ovaries retained in the fish and as tension changes in ovaries removed from the fish, suspended in a smooth muscle bath, and connected to a force transducer. Spontaneous contractions were observed in approximately one‐half the ovaries tested, using both methods. The occurrence of spontaneous contractions was dependent on the reproductive stage of the fish, with more sexually mature fish exhibiting a greater incidence of spontaneous activity. The rate of contraction increased linearly with temperature over the range 20–35°C. Although the physiological significance of ovarian contractions in F. heteroclitus is unclear, possible functions include the facilitation of ovulation, egg release during spawning, and the movement of eggs or fluid within the ovary. © 1992 Wiley‐Liss, Inc.

Ovarian muscularis of the stable fly Stomoxys calcitrans: Its structural, motile, and pharmacological properties

AbstractThe muscles of the ovary of the stable fly (Stomoxys calcitrans) appear to consist of two groups: (1) a network of stellate muscle cells that cover the surface of the ovary and (2) fibers that surround the ovarioles. Innervation of the ovaries was largely restricted to the region of the pedicels. The structural arrangements of the ovarian muscles provided the basis for two distinct patterns of movement. The contraction of the sheath that surrounds the ovary produced the appearance of a pulsing sphere, while the activation of muscle fibers that encompass the ovarioles may cause a vertical translation of eggs within the tubes. The various patterns of motility derived from the combined and separate actions of these two groups of muscles are described. Oscillations in the size of the ovary were the most prominent and frequent kind of spontaneous activity observed. A complete cycle of oscillation ranged from 200 ms to 4 s. Day‐old stable flies consistently had the highest rate (30%) of ovarian sheath compression of the three age groups examined, while 2‐day‐old flies had the lowest (less than 10%). Seven‐day flies had inconsistent rates that ranged from o to 25%. Two kinds of ovarian compression were recognized on impedance myographs on the basis of small and large amplitudes. Changes in tonus were also detected. Octopamine produced large increases in the amplitude of spherical compressions at 10−7 M. The neuropeptide proctolin (10−9 M) caused changes in both the frequency and amplitude of ovarian contractions. Glutamic acid consistently caused a marked reduction in the amplitude of ovarian compressions.

The autonomic innervation of the ovary of the dab,Limanda yokohamae

The autonomic innervation of the ovary of the dab was studied histologically and physiologically. The ovary receives a branch of nerve bundles that emerge into the abdominal cavity at the postero-ventral end of the kidney and can be traced back to the sympathetic chain in the vicinity of the 5th vertebra. Almost all the nerve fibers are AChE-positive, and some of them also emit adrenergic fluorescence. Electrical stimulation of the ovarian nerves caused ovarian contractions, and administration of ACh elicited contractions of the ovary preparations, supporting the hypothesis that the ovary is innervated by excitatory cholinergic fibers. In the ovarian nerve bundles, many AChE-positive and non-fluorescent ganglion cells are scattered. Ultrastructural studies suggest that nerve endings situated on the ovarian smooth muscle and on ganglion cells are cholinergic. These results also suggest that the cells are the post-ganglionic neurons of the cholinergic innervation and the axons of the cells reach to the muscle cells. On the other hand, the adrenergic fluoresecent fibers possibly participate in the inhibitory innervation, since the presence of inhibitory beta-adrenoceptors were demonstrated by pharmacological studies.

Effects of drugs on the responses of the ovary to field and nerve stimulation in a tilapia Sarotherodon niloticus.

The effects of drugs on the tonus changes of ovarian smooth muscles of tilapia Sarotherodon niloticus were studied on isolated preparations. Excitatory action of acetylcholine was abolished by atropine and enhanced by neostigmine, indicating the presence of muscarinic acetylcholine receptors on smooth muscle cells. The cholinergic contractions of the ovary were inhibited by the beta adrenergic action of sympathomimetic amines (isoprenaline, adrenaline and noradrenaline), since their inhibitory action was abolished by a beta adrenoceptor antagonist, propranolol. After an application of propranolol, the excitatory alpha adrenergic action of the amines, except isoprenaline, was uncovered, indicating that alpha adrenoceptors are also located on them. Both field and paravascular nerve stimulations activated cholinergic excitatory nerves and caused ovarian contractions, since the contraction was blocked by atropine and tetrodotoxin. From the present results, excitatory cholinergic innervation of the tilapia ovary was confirmed. However, no conclusive evidence to show inhibitory adrenergic innervation was obtained.

The fate of thecal smooth muscle cells in postovulatory hamster follicles.

Smooth muscles cells (SMC) have been identified in the theca externa of preovulatory follicles in several mammals. The function of these cells is not clear. It has been suggested that they are involved in ovulation and may play a role in the collapse of ruptured follicles and their transformation into corpora lutea (CL). In this study we used transmission electron microscopy (TEM) to examine the ultrastructure of SMC at the base of hamster follicles during the formation and regression of CL. For the first three days following ovulation, the CL are surrounded by a distinct theca externa composed of 1-3 layers of spindle-shaped cells. The cells were identified as SMC by the masses of filaments in the cytoplasm, dense attachment plaques and caveolae along the plasma membrane and an external lamina surrounding each cell. On the fourth day after ovulation, the theca externa is fragmented and TEM observations suggest the following three fates for theca externa cells: 1) many cells degenerate, 2) some cells persist as SMC, 3) some cells possess few or no filaments and appear similar to fibroblasts. The significance of these results is discussed in regard to ovarian contractions, the fate of CL, and the recycling of ovarian tissues.

Ovarian contractility and its relationship to ovulation: a review.

During the past decade there has been a renewal of interest in the contention that ovarian contractions are necessary for mammalian ovulation. The century-long deliberation on this controversial issue has been revived by current evidence that: I) muscle-like cells are present in the vicinity of mature ovarian follicles: 2) contractility is a basic property of ovarian tissue; 3) autonomic nerves extend into the mammalian ovary and 4) neurohumoral agents, prostaglandins and other metabolic substances present in ovulatory tissue can stimulate ovarian contractility. However, despite the extensive new data, it remains unclear whether ovarian contractions have an essential role in the ovulatory process. The problem is no longer a matter of insufficient information as considerable evidence is now available on the topic. What is needed is a comprehensive analysis of the existing data. This paper critically surveys the current information, in an effort to clarify the functional significance of ovarian contractility in the mechanism of ovulation. The review concentrates on studies that have been carried out during recent years. For a summary of the earlier work on ovarian “smooth muscle,” the interested reader is referred to the perceptive report by Kraus (1947). In addition, there are a number of recent papers which discuss various aspects of the subject

Lack of spontaneous ovarian contractions in the baboon

Histological observations on the ovaries of various species reveal smooth muscle fibres throughout the stroma and theca interna (Lambertsen, Greenbaum, Wright & Wallach, 1976). Ovarian contractions have been observed and recorded in vitro and in vivo in an equally diverse number of species (Wright, Wallach, Fromm & Jeutter, 1976). Although the physiological significance of spontaneous ovarian contractions is not known it has been suggested that they may be in some way related to the mechanism of ovulation (Lipner & Maxwell, 1960). This speculation is supported by the observation that the frequency of spontaneous ovarian contractions increases about the time of ovulation (Virutamasen, Wright & Wallach, 1973). The methods which have been used to monitor ovarian contractions in vivo are not capable of clearly delineating contractions of ovarian origin from ovarian movement resulting from contrac¬ tions of the supporting mesenteries and ligaments of the ovary. Recent work in our laboratory has led to the development of very small doughnut-shaped carbon-impregnated silicone-rubber (Silastic S-2086, Dow Corning) transducers which are highly sensitive to direct contractile activity and rela¬ tively insensitive to movement. Instrumentation which provides for the biotelemetric transmission of the signal from these transducers permits their use in unrestrained, unanaesthetized animals for long periods of time. Moreover, because of the small size of the sensor (approximately 1200 pm in diameter), recording of contractile activity can be made at a precise location of interest without sig¬ nificant interference from movement of the structure being monitored. The method of making and calibrating the transducers and a description of the dual-channel biotelemetric system used for signal processing has been previously described (Blair & Beck, 1976; Blair, Gilliland & Sauer, 1976). To test the question of whether or not ovarian tissues of the baboon (Papio papio) undergo spontaneous contractions, transducers of this type were positioned within the ovarian stroma of 3 adult baboons and observations were made on the contractile activity over a 30-day period. All 3 females were experiencing normal menstrual cycles. Two transducers were placed in each animal :

The influence of ovarian denervation and nerve stimulation on ovarian contractions

Two techniques were used to study the neuromuscular control of rabbit ovarian contractions. First, the ovary was denervated by stripping its artery. Spontaneous contractions were noted, as was a normal response to exogenously administered adrenergic agents. Second, the ovarian nerves were electrically stimulated in an in vitro perfused ovary system. Inhibition of ovarian contractions was noted, and the additive effects with adrenergic agents suggested a betamimetic effect of nerve stimulation in the estrous rabbit. After administration of exogenous human chorionic gonadotropin, the response was reversed. The significance of the contribution of a neuromuscular mechanism to the control of ovulation is discussed.

Studies of Rabbit Ovarian Contractility Using Chronically Implanted Transducers

A chronically implanted, highly sensitive force transducer was used to study ovarian contractions in the rabbit. The transducer is implanted into the medulla of the ovary through its long axis. The leads are then drawn through the abdominal wall and directed subcutaneously to the back of the neck. The miniature pin connector is fixed into place beneath the skin to facilitate accessibility for repeated recordings, obviating the necessity for anesthesia or serial laparotomies. Intraovarian transducers were implanted in isolated rabbits. Serial recordings of ovarian contractions were made at weekly intervals for 3 weeks, following which ovulation was induced with 100 IU of human chorionic gonadotropin (HCG). Extended recordings were made from 3 through 13 hours after HCG administration to six animals. Recordings revealed increased ovarian contractile activity beginning just before the anticipated time of ovulation. The observations support those of previous acute experiments and demonstrate a relationship between ovarian contractile activity and the process of follicle rupture.

Spontaneous and Prostaglandin- or Oxytocin-Induced Motility of Rat Ovaries Isolated During Different Stages of the Estrous Cycle: Effects of Norepinephrine

Ovaries isolated from rats in different stages of the sex cycle were explored for spontaneous or drug-induced contractile activity. The number of spontaneously active ovaries as well as the magnitude of the isometrically developed tension and frequency of contractions were greater during the periovulatory interval (late proestrus and estrus) than during early proestrus or metestrus. Furthermore, during estrus or late proestrus the left ovaries exhibited a mechanical activity significantly greater than that of the right ovaries. The oxytocin-triggered motility was clearly more marked immediately prior to ovulation (late proestrus) and greater in left ovaries than in right ovaries. In contrast, the contractions induced by prostaglandin F2alpha were similar during early proestrus and late proestrus. Ovarian contractile reactivity to norepinephrine indicated the presence in the tissue of alpha- and beta-adrenergic receptors. During early proestrus this agent stimulated the motility of left and right ovaries, whereas close to the ovulatory interval (late proestrus) it depressed the contractions of left ovaries. This last influence was blocked by propranolol. The existence of a close relationship between ovarian contractile activity and ovulation is reinforced by the present results in the rat. A tentative participation of oxytocin is also suggested. In addition, the influences of other possible regulatory agents of ovarian contraction, such as catecholamines and prostaglandins, are presented and discussed.

Intraovarian Pressure Changes During Ovulation in Rabbits

Contractile elements are found in the ovaries of many species, but it has not been possible to ascertain whether these elements are of importance in the process of ovulation. In this report, we describe changes in intraovarian pressure recorded continuously in vivo in unanesthetized rabbits under normal conditions and under the influence of intravenously injected human chorionic gonadotropin (hCG), as well as following the ovulatory stimulus of normal copulation. The recordings were made by means of small latex balloons (0.02- to 0.04-ml volume) attached to indwelling catheters, inserted into the ovarian stroma, and secured with 6-0 nylon sutures. All 24 rabbits studied showed changes in intraovarian pressure indicative of ovarian contractile activity. The intraovarian pressure changes followed a characteristic pattern which was different from the changes in intratubal pressure, recorded simultaneously from the lumen of the ipsilateral fallopian tube, indicating that the contractions of both organs occurred independently. In normal animals, before an ovulatory stimulus was applied, the ovarian contractility pattern consisted of a series of rapid contractions (average amplitude, 6 mm Hg; average frequency; 8 per minute) occurring with intervals of quiescence lasting from 11 to 36 minutes. The base line tonus was frequently elevated during these series of contractions. Mating or an injection of hCG had no immediate effect on intraovarian pressure but, 6 to 8 hours after the stimulus was applied, ovarian contractile activity increased significantly in all rabbits. This enhanced activity persisted for several hours before returning to initial levels approximately 15 to 18 hours after mating or the hCG injection. This demonstration of increased contractile activity about the time of ovulation suggests that ovarian contractions participate in the process of follicular rupture and the extrusion of ova at ovulation. Prostaglandin F2alpha, norepinephrine, and oxytocin were effective in inducing ovarian contractions.

Effects of indomethacin and prostaglandin F 2α on ovulation and ovarian contractility in the rabbit

The present investigation was carried out to determine whether inhibition of ovulation in the rabbit by administration of indomethacin can be correlated with any change in ovarian contractility at ovulation time and can be reversed by administration of prostaglandins. Indomethacin was adminstered intra-muscularly using three different schedules in a dose of 5 mg/kg. A reduced number of ruptured follicles following HCG was noted in all groups treated with indomethacin. Infusion of PGF 2α into the aorta (1 μg/kg/min.) could reverse this effect. Less pronounced ovarian contractility was observed after indomethacin treatment, but infusion of PGF 2α immediately enhanced contractility in ovaries from indomethacin treated rabbits. The inhibition of ovulation in the rabbit associated with indomethacin adminstration may be related to suppression of ovarian contractions. These data also suggest that prostaglandins may play a significant role in the mechanism of ovulation through an influence on ovarian contractility.

Observations of spontaneous contractions of the cat ovary in vitro.

Cat ovaries were isolated in an organ bath containing physiological solution. Isometric contractions of the ovary were recorded. At least one ovary in 80% of the normal cats showed spontaneous rhythmic contractions. Contractions were also observed in pregnant cat ovaries. In general, low doses of norepinephrine and epinephrine had a stimulatory effect on the frequency of spontaneous contractions, although a reduction in frequency of contractions was observed infrequently in the normal, pregnant and estrogen-treated ovaries. Isoproterenol caused a reduction or complete inhibition of frequency of spontaneous contractions. Phenoxybenzamine blocked the stimulatory effect of norepinephrine. Propranolol blocked the inhibitory effect of isoproterenol. It therefore appears that adrenergic receptors may be present. The physiological significance of the ovarian contractions is unknown. (Endocrinology84: 1336, 1969)